Project Summary ? PROJECT 2: JI, Gordon Bennett Insects play fundamental roles in human health by vectoring diseases and impacting food production. The success of insects is largely due to their symbiotic associations with beneficial bacteria that provide essential nutrition. This interaction has important parallels both with other animal-microbiome associations vital to host metabolism, and with host-pathogen interactions that rely on similar cellular mechanisms of nutritional exchange. However, bacterial microbiomes are diverse, have wide-ranging impacts on their hosts, and the cellular mechanisms that govern these interactions remain poorly understood. By developing a simplified model system, the proposed project will determine how animals regulate individual members of its microbiome at the cellular level to achieve balanced, essential nutrition. Preliminary work has identified a simple insect- bacteria symbiosis wherein the host relies on two bacterial species for nutrition. The proposed research will use a suite of advanced genomic, gene editing, and metabolomic approaches to (i) profile the genetic contributions of the host and its microbiome, (ii) determine how nutrients are synthesized, and (iii) understand how nutrients are exchanged between partners. Sequencing of host and bacterial genomes will provide a precise picture of the genetic determinants that underlie host-microbiome interactions. In addition, experiments will manipulate host diet to identify compensatory shifts in gene expression and metabolite production. Outcomes will indicate the genetic pathways involved in maintaining nutritional balance, and these conclusions will be experimentally verified with direct gene manipulations using CRISPR/Cas9.!By delineating how the host and its microbiome respond to diet, results will be leveraged to understand the cellular mechanisms that regulate and integrate the metabolisms of each partner. Developing this insect model, and associated research technologies, offers a critical approach by which to simplify issues that currently challenge direct investigation of host-microbiome interactions at the cellular level.